You have been given a blood sample from a patient with a bleeding disorder. You need to purify thrombin (a blood clotting enzyme present in blood serum) so that it can be characterized.
1) The first step in the purification is to separate serum proteins, including thrombin, from the whole cells present in the blood sample. What method could you use for this separation? What fraction of the resulting separation would you save?
2) You have managed to separate thrombin from all but two other serum proteins (see the table below). You run the proteins through an anion exchange column using a buffer at pH 6.4.
Protein pI Size (kDa) Cytochrome c 10.6 12.0 Lactoglobin 5.2 18.4 Thrombin 7.1 36.0
i) Which protein(s) remain(s) bound to the column?
ii) How could you elute thrombin and achieve proper separation?
PLEASE ANSWER ALL PARTS AND EXPLAIN IN DETAIL FOR POINTS! Thank you :)
You have been given a blood sample from a patient with a bleeding disorder. You need to purify thrombin (a blood clotting enzyme present in blood serum) so that it can be characterized.
1) The first step in the purification is to separate serum proteins, including thrombin, from the whole cells present in the blood sample. What method could you use for this separation? What fraction of the resulting separation would you save?
2) You have managed to separate thrombin from all but two other serum proteins (see the table below). You run the proteins through an anion exchange column using a buffer at pH 6.4.
| Protein | pI | Size (kDa) |
| Cytochrome c | 10.6 | 12.0 |
| Lactoglobin | 5.2 | 18.4 |
| Thrombin | 7.1 | 36.0 |
i) Which protein(s) remain(s) bound to the column?
ii) How could you elute thrombin and achieve proper separation?
PLEASE ANSWER ALL PARTS AND EXPLAIN IN DETAIL FOR POINTS! Thank you :)
